Use of stilbene compounds in preparing medicaments for treating or preventing diabetes and diseases associated with retrovirus

ABSTRACT

This invention relates to a new use of stilbene derivatives or pharmaceutically acceptable salts thereof, especially in the manufacture of medicament for the prevention and treatment of diabetes or retrovirus associated diseases.

FIELD OF THE INVENTION

This invention relates to a new use of stilbene derivatives orpharmaceutically acceptable salts thereof, especially in the manufactureof medicament for the prevention and treatment of diabetes or retrovirusassociated diseases.

TECHNOLOGY OF THE BACKGROUND

Diabetes is a common metabolic disorder in human beings. Recently, alongwith the improvement of living standard, and the changing of foodstuffstructure, the incidence of diabetes are increasing rapidly. In theworld there are around 0.12 billion of patients suffering from thisdisease. It is a serious threat to mankind. Therefore the prevention andtreatment of diabetes is a hot focus in the field of medicinal researchwork.

Now the anti-diabetic medicament used in clinics such as sulfanylureas,biguanidins etc are effective yet with some side effects. Someformulation derived from Chinese traditional herbs are effective, lesstoxic. Up to now no hypoglycemic monomer derived from natural plantswhich is used in clinics is reported.

OBJECTION OF THE INVENTION

Objection of this invention is to develop a new use of stilbenederivatives or pharmaceutically acceptable salts thereof.

SUMMARY OF THIS INVENTION

The investigation of the inventors has discovered that the stilbenederivatives of formula I or pharmaceutically acceptable salts thereofhave positive hypoglycemic effect and anti-retrovirus effect, then theycould be useful for prevention and treatment of diabetes andretrovirus-associated diseases.

Therefore, the first aspect of this invention relates to a use of atleast one stilbene derivatives of formula I or pharmaceuticallyacceptable salts thereof in the manufacture of medicament for theprevention and treatment of diabetes or retrovirus-associated diseases,

wherein, R₁, R₂, R₃ and R₄ are individually H, —OH, alkyl, C₆₋₁₀ arylsuch as phenyl or naphthalenyl, alkylhydroxyl- , alkoxyl and sugarcontaining glycosides such as —O-glucosyl or -glucosyl.

The second aspect of this invention relates to a composition for theprevention and treatment of diabetes or retrovirus-associated diseaseswhich comprising at least one stilbene derivatives of formula I orpharmaceutically acceptable salts thereof, pharmaceutically acceptablecarrier or excipient,

Wherein, R₁, R₂, R₃ and R₄ are independently H, —OH, alkyl, C₆₋₁₀ arylsuch as phenyl or naphthalenyl, alkylhydroxyl-, alkoxyl and sugarcontaining glycosides such as —O-glucosyl or -glucosyl.

Furthermore, this invention relates to a method of the prevention andtreatment of diabetes or retrovirus-associated diseases which comprisingadministrating a effective amount of stilbene derivatives of formula Ior pharmaceutically acceptable salts thereof to the patient.

wherein R₁, R₂, R₃ and R₄ are independently H, —OH, alkyl, C₆₋₁₀ arylsuch as phenyl or naphthalenyl, alkylhydroxyl-, alkoxyl and sugarcontaining glycosides such as —O-glucosyl or -glucosyl.

DETAILED DESCRIPTION OF THIS INVENTION

According to this invention, the compound derivatives of formula I maybe obtained from a natural plants or by a chemical synthesis. Forexample, said compounds of formula I could be extracted from the naturalplant such as Vitis L, Ampelopsis Michx of Vitaceae; Arachis L, CassiaL, Sophora L, of Leguminosae; Veratrum L of Liliaceae; EucalyptosL'H'erit of Myrtaceae; and Rheum emodi Wall, Rheum franzenbachii Munt,Rheum hotaoense C. Y. Chang, Rheum wittrockii Lundstr, Rhizoma polyonicuspidati of Polysonaceae According to this invention, the term“diabetes” used in this invention means to type I and for type IIdiabetes.

According to this invention, the term “patient” in this inventiondenotes mammalians such as human beings.

According to this invention, the term “alkyl group” denotes a loweralkyl containing 1-6 carbon atoms, the alkyl in terms “alkylhydroxylgroup” or “alkoxyl” is defined as above definition of alkyl.

According to this invention, the retrovirus-associated diseases denotehepatitis or HIV infected diseases.

According to this invention, the preferred compounds of formula I inthis invention are selected from:

-   -   3,4,5-trihydroxystilbene (compound E),    -   3,3′,4′,5-quadrahydroxystilbene-4′-O-β-D-glucopyranoside        (Compound E₁)    -   3,4′,5-trihydroxy-3′-methyloxy stilbene-3-O-β-D glucoside        (compound E₂)    -   3,5-dihydroxy-4′-methyloxy stilbene-3-O-β-O-D glucoside        (compound E₃)    -   3,4′,5-trihydroxy stilbene-3′-O-D-glucoside (compound E₅)

According to this invention, stilbene derivatives of formula I may beformulated into to enteric or parenteral dosage forms such as tablet,capsule, granule or injection etc, by the known manner in the art.

Following examples will further illustrate this invention in detail butdo not represent any limitation to the scope of the invention.

EXAMPLE 1 Preparation of 3,4,5-trihydroxystilbene (compound E)

Pour 95% alcohol to Huchan slices in proportion of 8:1 (V/W) was mixedand the obtained mixture was extracted for 3 cycles, 2 hrs per cycle,combining the extract solution, concentrating it in low pressurecondition. The concentrated extract was dispersed by water, degreasingwith ether, then extracting with ethyl acetate, n-butyl alcohol. Theethyl acetate fraction, n-butyl alcohol fraction and water fraction wereobtained respectively. Separating ethyl acetate fraction on silica gelcolumn (mash 60-100), compound E crude product was collected by ethylacetate-methyl alcohol gradient elution, then re-crystallizing withacetone. N-butyl alcohol fraction was separated on silica columnchromatography, with ethyl acetate gradient elution, compound E₅ productwas collected, and re-crystallizing with acetone-water.

Identification:

Compound E is white needle crystal m.p.253-255° C., easily soluble inmethyl alcohol, ethyl alcohol and acetone etc. FeCl₃ reaction showsgreen in color. Blue to violet fluorescence is excited by UV light.

Uvλ max MeOH (nm): 216,303.

IR(KBr)cm⁻¹: 3240,1880,1585,965.

¹HNMR(acetone-d₆)δ ppm:8.79(1H,Br.s,4′-OH),8.48(2H,Br.s,3,5-H),7.36(2H,dd,J=2.4/8.5Hz,H-2′,6′), 6.95(1H,d,J=16.2 Hz,H-β),6.81(1H,d,J=16.2 Hz,H-α),6.78(2H,dd,J=2.4/8.5 Hz,H-3′5′),6.77(2H, d,J=2.2 Hz,H-2,6),6.48(1H,t,J=2.4Hz,H-4).

¹³CNMR(acetone-d₆)δ ppm: 159.47 (C-3,5),158.08(C-4′),140.73(C-1),140.73(C-1),129.78(C-1′),128.98(C-2′,6′),128.60(C-α),126.74(C-β), 116.29(C-3′,5′),105.47(C-2,6),102.51(C-4).EI-MS

m/z: 228(M⁺,100),227(M⁺-1),211(M⁺-OH),181,157,115,91,76.

Spectrum data is reported by Ming Te et al(1.Ming Te et al: Journal ofChinese traditional Medicine 1998,28(8):486) Therefore compound E isidentified as 3,4,5-trihydroxystilbene or resveratrol.

EXAMPLE 2 Preparation of 3,3′,4′,5-quadrahydroxystilbene-4′-O-β-D-glucopyranoside (compound E₁)

Pour 95% ethyl alcohol to root and rhizome of Rheum emodi Wall inproportion of 8:1(V/W) was mixed, refluxed for 3 cycles, 2hr per cycle,combining the extract solution, concentrating in low pressure condition.The alcohol extract was dispersed by diatomite and drying.

Washing off lipid soluble fraction with chloroformn. Further elutionwith ethyl acetate, collecting soluble fraction. Separating thisfraction by elution with ethyl acetate on silica gel chromatography(mash 60-100), then eluted with ethyl acetate/methanol (4:1-2), E₁compound crude product was collected, and re-crystallizing withwater-acetone.

Identification

Compound E₁ is white amorphous powder (diluted acetone)

m.p.138-140° C.

Blue to violet fluorescence was excited by UV light. Molisch reactionwas positive.

¹HNMR(acetone-d₆)δ ppm: 7.14(1H,d,J=805 Hz,H-5′), 7.06(1 H,d,J=2.1Hz,H-2′), 6.97(1H,d,J=16.3 Hz,H-β), 6.94(1H,dd,J=2.1/8.5 Hz,H-6′),6.89(1H,d,J=16.3Hz, H-α),6.52(2H,d,J=2.1 Hz,H-2,6),6.24(1H,t,J=2.1Hz,H-4),4.79(1H,d,J=7.5 Hz,anome ric-H),3.9-3.3(sugar-H);

¹³CNMR(acetone-d₆)δ ppm: aglycone 159.5(C-3,5),148.5(C-4′),146.0(C-3′),140.3(C-1),134.2(C- 1′),128.6(C-α,β), 119.3 (C-5′),118.9(C-6′), 114.2(C-2′),115.6(C-2,6),104.0(C-4),glucosyl: 102.9(C-1″),77.8(C-3″),77.1(C-5″),74.4(C-2″),70.9(C-4″),62.2(C-6″).

The 1HNMR and 13CNMR data are reported by Yoshiki Kashiwada et al (2Yoshiki Kashiwada et al: Chem. Pharm Bull 1988,36(4):1545). Compound E₁is identified as piceatannol-4′-O-β-D-glucopyranoside

Example 3 Preparation of 3,4′,5-trihydroxy-3′-methyloxy stilbene-3-O-β-Dglucoside (compound E₂)

Compound E₂ was obtained from root and rhizome of Rheum hotaoense C. Y.Chang by the same procedure as to that in example 1 or example 2.

Identification:

Compound E₂ is while needle crystal (diluted methyl alcohol)

m.p.228-230° C.

Blue to violet fluorescence is excited by UV light. Molisch reaction ispositive.

¹HNMR (acetone-d₆)δ ppm: 7.07(1H,d,J=2.0 Hz,H-2′), 7.02(1H,d,J=16.5Hz,H-β),6.96(1H,dd,J=2.0/8.3 Hz,H-6′), 6.90(1H,d,J=7.9 Hz,H-5′),6.89(1H,d,H=16.5Hz,H-α),6.77(1H,Br.s,H-2),6.66(1H,Br.s,H-2),6.48(1H,t,J=1.8Hz,H-4),4.90(1H,d,J=7.7 Hz,anomeric-H),3.82(3H,s,—OCH₃),4.0-3.3(sugar-H);

¹³CNMR(acetone-d₆)δ ppm: aglycone 160.1 (C-5),159.5(C-3),148.4(C-4′),147.5(C-3′),140.5(C-1),131.5(C-1′),129.5(C-β),127.2(C-α),119.7 (C-6′), 113.3(C-2′),112.5(C-5′),108.0(C-2),106.5(C-6),103.8(C-4),56.2(—OCH₃);glucosyl:101.9(C-1″), 77.7(C-3″,5″),74.4(C-2″),71.1(C-4″),62.5(C-6″).

1HNMR and 13CNMR data are reported by Yoshiki Kashiwada et al (3 YoshikiKashiwada et al: Chem. Pharm Bull 1984.32(9): 3501), Compound E₂ was

identified as 3,4′,5-trihydroxy-3′-methyloxy stilbene-3-O-β-D-glucoside,or rhaponticin)

E₂ 3,4′,5-trihydroxy-3′-methyloxy stilbene-3-O-β-D-glucoside, orrhaponticin EXAMPLE 4 Preparation of 3,5-dihydroxy-4′-methyloxystilbene-3-O-β-O-D glucoside (compound E₃) and 3,4′,5-trihydroxystilbene-3′-O-D-glucoside (compound E₅)

Compound E₃ or E₅ were obtained from root and rhizome of Rheumfranzenbachii Munt or Rhizoma polygoni cuspidati by almost the sameprocedures as those in example 2.

Identification:

Compound E₃ is colorless needle crystal (acetone),

m.p.210° C.,

Blue to violet fluorescence is excited by UV light. Molisch reaction ispositive.

Uv λ max MeOH (nm): 216,296.IR(KBr)cm⁻: 3455,3320(OH),1595,1505,830,772,675.

¹HNMR(acetone-d₆)δ ppm: 7.51(2H,d,J=8.6 Hz,H-2′,6′),7.08(1H,d,J=16.6Hz,H-α), 6.94(1H,d,J=16.6 Hz,H-β),6.91(1H,d,J=8.6Hz,H-3′,5′),6.70(2H,Br.s,H=2,6),6.35 (1H,t,J=2.2 Hz,H-4),4.81(2H,d,J=7.6 Hz,anomeric-H),3.76(3H,s,OCH₃),3.3-3.9(6H, m,sugar-H);

³CNMR(acetone-d₆)δ ppm: 159.2(C-5),158.5(C-3),159.0(C-4′),139.0(C-1),129.2(C-1′),128.0(C-β),127.3(C-2′,6′),126.0(C-α),114.5(C-3′5′),107.2(C-6),104.8(C-2),103.0(C-4)55.2(OCH₃),glucosyl:101.8(C-1″),74.0(C-2″),77.2(C-3″),70.8(C-4″),76.8(C-5″),61.8(C-6″).FAB-MS m/z: 404(M⁺),242(M⁺-glu).

Above data is reported by Ming De et al (1 Ming De et al: Journal ofChinese traditional medicinal herbs,1998,23 (8):486). Therefore E₃ isidentified as 3,5-dihydroxy-4′-methyloxy stilbene-3-O-β-D-glucoside, or(desoxyrhaponticin)

3,5-dihydroxy-4′-methyloxy stilbene-3-O-β-D-glucoside, ordesoxyrhaponticin

Compound E₅ is white long needle crystal (acetone-water)

m.p.228-230° C., easily soluble in acetone.

Blue to violet fluorescence is excited by UV light. Molisch reaction ispositive. Uv λ max MeOH (nm): 220,303.

IR(KBr)cm¹:3610,3310,2975,2923,2880,1610,1589,1516,1450,1360,1320,1250,1170,1075,965,840. ¹HNMR(acetone-d₆)δ ppm:8.89(1H,Br.s,4′-OH),8.86(1H,Br.s,5-OH),7.35(2H,dd,J=2.4/8.5 Hz,H-2′,6′),7.20(1H,d,J=16.2 Hz, H-β),6.84(1H,d,J=16.2 Hz,H-α),6.78(2H,dd,J=2.4/8.5Hz,H-3′5′),6.73(1H,Br.s,H-β),6.62(1H,Br.s,H-2),6.45(1H,Br.s,H-4),4.88(1H,d,J=7.7Hz,anomeric-H),3.8-3.2 (sugar-H).

³CNMR(acetone-d6)δ ppm: 160.10(C-3),159.33(C-5),158.19(C-4′),140.73(C-1),129.64(C-1′), 129.56(C-2′,6′),128.70(C-α),126.33(C-β),116.31(C-3′,5′), 108.00(C-2),106.49(C-6),103.72(C-4),glucosyl:101.90(C-1″),74.57(C-2″),77.76(C-3″),71.26(C-4″),77.64(C-5″),62.48(C-6″).FAB-MS m/z: 389(M⁺-H),242(M⁺-glu). .EI-MS m/z:228(M⁺,100),227(M⁺-1),211(M⁺-OH),181,157,115,91,76.

The spectrum data is reported by Wang ZhenYu et al(4 Wang ZhenYu et al:Chinese herbs 1996,27(12): 714) Therefore E₅ is identified as3,4′,5-trihydroxy stilbene-3′-O-β-D-glucoside or ploydatin.

3,4′,5-trihydroxy stilbene-3′-O-β-D-glucoside or ploydatin. EXAMPLE 5Biological Activity Assay

Following biological experiment demonstrated that the effect of thecompounds in examples 1-5 on glucose tolerance curve of normal mice oron glucose levels of alloxan-induced diabetic mice. The metformin ormiglucan were used as positive control drugs. And the hypoglycemiceffects were evaluated. Hypoglycemic effect of stilbene derivatives onglucose level of alloxan induced diabetic mice.

KM strain male mice which have been fasted for 5-8 hours are used and beinjected intravenously of alloxan 80mg/Kg, and 72 hours later, selectedthose with glucose level>11.0 mmol/L as the diabetic model mice for drugevaluation. Compounds were administered pos for 12 days. The results areshown in table 1: TABLE 1 Hypoglycemic effect on glucose level ofalloxan induced diabetic mice Mice Dosage Predosing Postdosing CompoundGroup (no) (mg/kg) (mmol/L) (mmol/L) E Normal 11 H₂O  6.91 ± 1.01  7.31± 0.80 Model 11 H₂O 17.30 ± 4.57 25.23 ± 9.17 Metformin 10  500 17.10 ±4.84 15.16 ± 8.53 E L 11  150 17.87 ± 4.86 18.36 ± 6.25 E S 11  75 17.27± 4.60 20.01 ± 7.1  E₁ Normal 11 H₂O  7.14 ± 1.18  7.07 ± 1.32 Model 12H₂O 22.89 ± 3.94 22.95 ± 2.94 Metformin 13  500 22.81 ± 4.89 17.49 ±5.33 E₁ L 13 1000 22.65 ± 3.55 16.54 ± 5.48 E₁ M 13  500 22.76 ± 3.1716.39 ± 5.32 E₁ S 13  250 22.49 ± 4.63 21.10 ± 5.73 E₂ Normal 8 H₂O 7.12 ± 1.15  6.38 ± 1.53 Model 8 H₂O 22.70 ± 4.33 27.79 ± 7.98Metformin 8  500 21.15 ± 3.85 17.64 ± 1.00 E₂ L 8 1000 21.38 ± 5.7524.64 ± 2.49 E₂ M 8  500 22.98 ± 5.94 28.96 ± 8.21 E₂ S 8  250 21.43 ±5.04 22.61 ± 6.07 E₅ Normal 11 H₂O  6.91 ± 1.01  7.31 ± 0.80 Model 11H₂O 17.30 ± 4.57 25.23 ± 9.17 Metformin 10  500 17.10 ± 4.84 15.16 ±8.53 E₅ L 11  150 17.92 ± 5.35 21.05 ± 4.63 E₅ S 11  75 17.46 ± 5.8621.79 ± 9.39Table 1 indicated that metoformin, positive control drug, is effectivein the experiments, and the compounds of this invention E, E₁-E₅ areeffective too, although some with higher or lower efficacy.Effect of the Compounds of Examples 1-5 on Glucose Tolerance Curve ofNormal Mice or Rats

In these experiments, stilbene derivatives in examples 1-5 areadministrated respectively to KM male mice or Wistar male rats pos for12 days , then test animals were fasted for 8 hours, examined glucosevalues, and administrated the test compound. One hour later,administered ip of glucose 2 g/Kg (1.11 mol/L glucose solution).Determining the glucose values at 0, ½, 1, 2 hours after glucoseinjection and calculating the area under this glucose tolerance curve.The results are shown in table 2: TABLE 2 Effect of E and E5 on glucosetolerance of normal rats (N = 6) Dosage Glucose (mg/ (fasted) Glucoseafter medication (mmol/L) Group Kg) (mmol/L) 0′ 30′ 60′ 120′ Auc Normal/ 5.4 ± 0.4 6.0 ± 0.8 12.5 ± 2.3 8.5 ± 1.7 6.6 ± 0.9 1044.2 ± 141.8 Miglucan 100 5.3 ± 0.7 3.4 ± 0.4  8.7 ± 3.5 5.3 ± 2.1 3.8 ± 1.4 663.8 ±220.2 E L 150 5.8 ± 0.6 4.2 ± 0.4  9.5 ± 2.1 6.1 ± 1.2 4.7 ± 0.7 760.2 ±119.6 E S  75 5.3 ± 0.9 4.9 ± 0.7 12.1 ± 2.3 7.5 ± 0.8 6.2 ± 0.9 961.1 ±115.9 Normal / 4.7 ± 0.8 5.2 ± 1.4  9.7 ± 0.5 6.5 ± 0.9 5.8 ± 0.8 853.1± 87.1  Miglucan 100 2.2 ± 0.2 2.7 ± 0.8 11.5 ± 0.6 7.8 ± 6.2 4.1 ± 1.7883.1 ± 43.5  E₅ L 150 4.7 ± 0.8 4.3 ± 0.9 10.2 ± 1.5 6.3 ± 1.1 5.7 ±1.1 836.3 ± 112.1 E₅ S  75 4.6 ± 0.5 4.9 ± 0.7 12.0 ± 1.5 7.4 ± 0.8 6.7± 0.7 966.8 ± 71.6 

Table 2 indicated that compounds of this invention effectivelystimulated the secretion of insulin in case of glucose loading. It issuggested that they can be useful for the treatment or prevention oftype II diabetes

Toxicity of compounds of this invention is shown in table 3. TABLE 3Maximal tolerance dose of the compounds of this invention in mice (pos)Compound Sex of mice MTD(g/kg) E ♀ >6.10 ♂ >6.00 E₁ ♀ 5.625 ♂ 4.219 E₂♀ >16.872 ♂ >18.301 E₃ ♀ >11.200 ♂ >11.000 E₅ ♀ >10.00 ♂ >10.00Table 3 indicated that the toxicity of compounds E, E₁, E₂, E₃ and E₅ isvery low. According to acute toxicity classification proposed by WHO in1977, they could be classified as low toxic or even no toxic agents.

And they are much less toxic than Metformin or miglucan used in clinicsnow.

Inhibition of Compounds of This Invention on HBeAg Expression

Experimental method: use 10% DMEM(with G418 380 μg/ml)to cultivate2.2.15 cells,

Add 1.5ml of 10⁵ cell/ml to 24 well plate for cultivation, change thenew cultivate fluid next day and add different amount of compound to it,cultivate each concentration of compound to 3 wells, then collect andfreeze 200 μl of supernatant of 2.2.15 cell culture at day 2,4,6cultivation. Measure HBeAg value of the supernatant by ELISA. Theresults are shown in table 4: TABLE 4 Inhibition of the compounds ofthis invention on expression of HBeAg 2.2.15 Cell(10⁵/ml) Day 2 Day 4Day 6 Compound OD IR (%) OD IR (%) OD IR (%) E₁₋₂ 0.244 57.75 0.20832.68 0.228 51.57 E₁₋₃ 0.224 63.83 0.354 −62.75 0.235 48.82 E₂₋₂ 0.19971.43 0.188 45.75 0.251 56.69 E₂₋₃ 0.301 40.43 0.244 9.15 0.347 4.72E₃₋₂ 0.194 72.95 0.197 39.87 0.177 71.65 E₃₋₃ 0.265 51.37 0.256 1.310.236 48.43 2.2.15 Ctr 0.434 0.258 0.359# E₁₋₂ and E₁₋₃ indicate compound E in concentration of 10 μg/ml and 20μg/ml, E₂.E₃ are the same, OD optical density, IR inhibition rate, ctr controlIR=[Ctr(P/N)−Agent(P/N)]/[Ctr(P/N)−2.1]Table 4 indicated that above compounds have inhibition activity on HBeAgexpression.

1. A use of at least one stilbene derivatives of formula I orpharmaceutically acceptable salts thereof in the manufacture ofmedicament for the prevention and treatment of diabetes orretrovirus-associated diseases,

wherein R₁, R₂, R₃ and R₄ be independently H, —OH, alkyl, aryl suchphenyl or naphthalenyl alkylhydroxyl-, alkoxyl and sugar containingglycosides such as —O-glucosyl or -glucosyl.
 2. A composition for theprevention or treatment of diabetes or retrovirus-associated diseaseswhich comprising at least one stilbene derivatives of formula I orpharmaceutically acceptable salts thereof, pharmaceutically acceptablecarrier or excipient,

wherein R₁, R₂, R₃ and R₄ be independently H, —OH, alkyl C₆₋₁₀ aryl suchas phenyl or naphthalenyl, alkylhydroxyl-, alkoxyl and sugar containingglycosides such as —O-glucosyl or -glucosyl.
 3. A method for theprevention or treatment of diabetes or retrovirus D-associated diseaseswhich including administrating effective amount of stilbene derivativesof formula I or pharmaceutically acceptable salts thereof to thepatients,

wherein R₁, R₂, R₃ and R₄ be independently H, —OH, alkyl C₆₋₁₀ aryl suchas phenyl or naphthalenyl, alkylhydroxyl-, alkoxyl and sugar containingglycosides such as —O-glucosyl or -glucosyl.
 4. Use as claimed in claim1, wherein said compounds are selected from: 3,4,5-trihydroxystilbene(compound E), 3,3′,4′,5-quadrahydroxystilbene -4′-O-β-D-glucopyranoside(Compound E₁) 3,4′,5-trihydroxy-3′-methyloxy stilbene-3-O-β-D glucoside(compound E₂) 3,5-dihydroxy-4′-methyloxy stilbene-3-O-β-O-D glucoside(compound E₃) 3,4′,5-trihydroxy stilbene-3′-O-D-glucoside (compound E₅)